The invention relates to the use of low-dose photodynamic therapy to modify donor tissue for xenograft transplantation. More particularly, it relates to exposing donor tissue intended for a recipient of a different species from the donor tissue to a photosensitizer and low intensity light.
PCT application publication no. WO96/21466 published Jul. 18, 1996, and incorporated herein by reference, describes a method for reducing the rejection of allografts by subjecting the donor tissue intended for an allograft recipient to low-light level photodynamic therapy. Skin allografts subjected to this treatment have enhanced survival times. Additional data demonstrating this effect are also described in U.S. Ser. No. 08/759,318 filed Dec. 2, 1996, and incorporated herein by reference. This work has also been reported by Obochi, M.O.K. et al., Transplantation (1997) 63:810-817.
These applications further mention that, in general, transplantation of tissue or organs is of three general types: syngeneic, where the donor tissue is of the same genotype as the recipient; allogeneic, wherein an allograft is derived from a donor of the same species as the recipient; and xenogeneic, where a xenograft is derived from a donor of a different species from the recipient. Transplantation using xenografts is of particular interest for use in human recipients since nonhuman donors can be used. However, model systems for such xenograft transplants can be constructed using two separate species of any derivations, such as rat/mouse, pig/bovine, and baboon/human.
Photodynamic therapy generally is a technique whereby a photosensitizer is administered directly to a tissue or organ or to a subject and the area desired to be treated is irradiated with light that is absorbed by the photosensitizer. In this state, the photosensitizer exerts an effect on the tissue or cells containing the photosensitizer. The effect may be cytotoxic or, alternatively, there may merely be an alteration in the function of the target cells or tissues. This approach has been used for selectively destroying tumor tissues, atherosclerotic plaques, the lesions of surface skin diseases, and unwanted pathogens in blood. Photodynamic therapy (PDT) has also been used to target activated cells of the immune system selectively.
Various effects of PDT on the immune system have been studied. It has been shown that photodynamic therapy using UV light, in particular 8-methoxypsoralen and UVA radiation, decreases 1 a antigens and the ATPase marker of epidermal Langerhans cells (LC) (Aberer, W. et al., J Invest Dermatol (1981) 76:202). The more standard photodynamic therapy combination of Photofrin(copyright) with visible light is able to inhibit APC from stimulating allogeneic cells in the MLR (Gruner, S. et al., Scand J Immunol (1985) 21:267). PDT with UVA light also inhibits upregulation of ICAM-1 expression by Langerhans cells (Tang, D. et al. J Immunol (1991) 146:3347). Photodynamic therapy using porphyrins inhibits the high affinity Fc-receptor on human monocytes (Krutmann, J. et al., J Biol Chem (1989) 264:11407).
Initially, PDT employed fairly high levels of radiation with the absorbed light. However, it has been shown that for certain applications, in particular modulating the immune system, only ambient light is necessary to activate the photosensitizer. Thus, xe2x80x9clow-levelxe2x80x9d PDT is conducted at much lower intensities of irradiation than those required to obtain a recognized photodynamic effect, such as skin erythema. A detailed description of low-dosage PDT and its effect on the immune system is found in PCT application publication no. WO96/22090 published Jul. 25, 1996, and in U.S. Ser. No. 08/856,921 filed May 16, 1997. In general, the radiation levels in xe2x80x9clow-dose PDTxe2x80x9d have upper limits in the range of 100 mW/cm2, more generally about 50 mW/cm2 or 25 mW/cm2 and lower intensities of less than 5 mW/cm2, preferably less than 1 mW/cm2 and more preferably less than 500 xcexcW/cm2 can also be used. Another criterion for radiation levels in low-dose PDT when an intact subject is administered low-dose PDT is that the light level used is less than one-quarter, preferably less than one-sixth, of that necessary to induce skin erythema in that subject. Still another measure of light dosage relates to total energy applied. Low-dose PDT employs energies of 10 J/cm2 or less. The above-referenced applications describing treating of donor tissue for allograft transplantation describe the use of low-dose PDT for that purpose.
Xenografts are generally more at risk for rejection than allografts. Success in regard to a protocol that diminishes the immunogenicity of allografts is clearly not predictive of the success of that technique in diminishing the immunogenicity of xenografts to the extent required to ensure acceptance by the recipient. For example, PCT application no. WO97/11653 published Apr. 3, 1997, purports to describe a protocol using standard photodynamic therapy at conventional light dosage levels to treat both allografts and xenografts to enhance their acceptability to recipient subjects. However, this technique is demonstrated to be useful, if at all, only with respect to allografts. The work described employs phthalocyanine, preinjects the donor animals with this drug, applies levels of 5 xcexc/ml of the drug after explantation and during irradiation and supplies radiation at 100 mW/cm2 for a total energy of 100 J/cm2. Fluence at this level kills the cells in the graft. The subject tissue is an aortic graft; indeed, the applicants point out that their technique is applicable only to grafts comprising an extracellular matrix and an amorphous ground substance.
It does not appear that the technique described in PCT application no. WO 97/11653 would thus successfully be applied either to xenografts or to graft tissue which comprises cell suspensions per se, such as those ordinarily used to treat Parkinson""s Disease. Other techniques known in the art also appear not to be sufficiently precise or successful to permit xenographic transplantation of neural tissue suspensions. Therefore, the source for replacement neural tissue in, for example, human patients with Parkinson""s Disease has been human fetal neural tissue.
On a wider scale, there is only limited experience with interspecies transplantation of organs into humans. The shortage of donors for human patients in need of liver, ocular tissue, cardiac tissue, lungs, and the like is well known. To alleviate this shortage, it has been suggested that animals of another species be used as sources for these donor materials. There have been several reports attempting to evaluate the possibility of using nonhuman sources for transplants into the central nervous system. See, for example, Pakzaban, P. et al., Neuroscience (1994) 62:989-1001 which surveys the literature regarding neural xenotransplantation and editorials by Sloan, D.J. et al. Neuroscience (1991) 14:341-346, Fishman, P. S. Neurobiol (1986) 36:389-391 and in The Economist (Mar. 22, 1997):99-101. An additional summary is that by Borlongan, C. V. et al., Neurological Res (1996) 18:297-304.
Fetal pig dopaminergic neurons have been transplanted into 12 patients with Parkinson""s; in one of these patients (who subsequently died), these neurons survived for at least 7 months (Deacon, T. et al., Nature Medicine (1997) 3:350-353). Attempts have also been made to modify the subject in whom the donor tissue will be implanted to become more receptive to the implant. For example, Honey, C. R. et aL, Exp Brain Res (1991) 85:149-152 treated recipient murine subjects with monoclonal antibody against the murine equivalent of CD-4 (L3T4) and then provided them with rat-derived POA grafts, resulting in longer survival times of the transplants. Cyclosporin-A is a standard method to immunosuppress recipients of xenografts. Pepino, P. et al., Eur Surg Res (1989) 21:105-113 used photochemotherapy and ciclosporin [sic] in recipient baboons of heterotrophic heart grafts from the cynomolgus monkeys (Macaca fascicularis).
Other treatments of recipient animals for xenogeneic transplantation have been reported, including treatment with antibody to interleukin-2 receptor (Honey, C. R. et al., Neuro Report (1991) 1:247-249), with antibodies to T cell receptors (Wood, M. J. et al., Neuroscience (1996) 70:755-789), with antibodies to T cellsper se (Okura, Y. et al., J Neurosci Res (1997) 48:385-396), with methyl prednisolone (Duan, W. et al., Brain Res (1996) 712:119-212), with FK506 (Sakai, K. et al., Brain Res (1991) 565:167-170), with 15-desoxyspergualin (Zhou, J. et al., Brain Res (1993) 621:155-160), and with 2-chlorodeoxyadenosine (Ryba, M. et al., Acta Neurobiologiae Experimentalis (1995) 55:259-270).
Thus, although PDT has been shown to inhibit the ability of lymphocytes to stimulate a mixed leukocyte reaction or to mediate graft host disease (Canti, G. et al., Photochem Photobiol (1981) 34:589) to inhibit the development of contact sensitivity in mice (Elmeths, C. A. et al., Cancer Res (1986) 46:1608; Simkin, G. et aL, Proc Opt Method Tumor Treatment Detect, SPIE (1995) 2392:23) and to inhibit rejection of skin allografts (Quin, B. et al., Transplantation (1993) 56:1481). There has been no suggestion that photodynamic therapy or low-dose PDT can be used to treat xenogeneic donor material for transplantation into a subject of another species.
Donor tissue has been treated with UV light (Reemtsma et al., U.S. Pat. No. 4,946,438; Lau et al., Science (1984) 223:607). It has been suggested that UVB radiation may inhibit LC antigen-presenting cell function by preventing the expression of critical co-stimulatory molecules (Simon et al., J Immunol (1991) 146:485). Several authors have suggested that the exposure of LC to UVB or psoralen plus UVA radiation (PUVA) causes a loss of surface markers (including ATPase and class II MHC antigens) without causing overt cytotoxicity (Aberer et al, (1981, supra); Hanau et al, J Invest Dermatol (1985) 85:135). However, Tang and Udey (Tang et al., J Invest Dermatol (1992) 99:83) showed that the levels of UV radiation that inhibited LC accessory cell function and selectively modulated ICAM-1 expression in short-term cultures were ultimately cytotoxic for LC.
Sometimes UV light has been used in conjunction with microencapsulation (Weber et al., U.S. Pat. No. 5,227,298). Other workers have used barrier membranes alone, such as the bilayer comprising a first non-cytotoxic layer and a second outer layer of a biocompatible and semipermeable polymeric material taught by Cochrum, U.S. Pat. No. 4,696,286.
Donor tissue has been treated with a wide variety of substances, such as the topical application of cyclosporin to skin grafts, as disclosed by Hewitt et al., U.S. Pat. No. 4,996,193, and the perfusion of a donor kidney with lymphocytic chalone, as described by Jones et al., U.S. Pat. No. 4,294,824. The survival time of skin grafts has been prolonged by treatment in vitro with cortisone, thalidomide, or urethane before implantation into a laboratory animal. The amount of drug locally applied to the skin is usually smaller than the amount required to achieve a similar effect by injecting the drug systemically into the recipient. Donor skin has been treated in vitro with streptokinase/streptodornase, RNA and DNA preparations of the recipient, or solutions of glutaraldehyde, prior to transplantation to reduce the antigenicity of the skin to be grafted.
More sophisticated approaches have involved the treatment of donor tissue with a monoclonal antibody directed against the MHC product along with complement (Faustman et aL, Proc Natl Acad Sci USA (1981) 78:5156) or the treatment of donor tissue with an immunoconjugate of antibody directed against the MHC (Shizuru, et al., Transplantation (1986) 42:660). Variable results were obtained by these methods.
For example, treatment of donor tissue has employed antibodies which mask the donor MHCI antigens (Pakzaban, P. et al., Neuroscience (1996) 65:983-996; Dismore, J. H. et al., Transplantation Proceedings (1996) 28:817-818). Extended time culturing of the donor tissue has also been suggested (Lafferty et al., Science (1975)188:259; Lafferty et al., Transplantation (1976)22:138-49; Bowen et al., Lancet (1979)2:585-86). Donor tissue has been treated with growth factor, such as TGF-beta (Czamiecki et al., U.S. Pat. No. 5,135,915), sometimes in combination with extended culture times (Orton, U.S. Pat. No. 5,192,312).
The use of PDT with xenografts appears to have been focused on exploration of PDT as a treatment for tumors. These studies employed in model systems, typically wherein human tumor tissue has been transplanted into immunodeficient mice. These studies do not relate to prolonging survival of the xenograft in the host. See, e.g., Gibson, S. L. et al. Brit J Cancer (1994) 69:473-481; Nelson, J. S. et al. J NCI (1988) 80:56-60; White, L. et al. Brit J Cancer (1988) 57:455-458; Makino, J Pediatric Surg (1986) 21:240-243; and Hill, J. H. et al. Am J Otolaryngol (1986) 7:17-27.
Despite the success of the use of low-dose photodynamic therapy in treating donor allografts, the enhanced difficulty of using cross-species donors for transplantation renders it surprising that this approach is also suitable for xenotransplantation. The difficulty in providing xenografts that will be accepted by the donor is well understood. While allograft tissue is clearly more at risk for rejection than syngeneic transplants, the quantum leap of species differences encountered when xenografts are substituted for allografts clearly distinguishes the behavior of these two types of donor tissues when ultimately transplanted into the recipient.
The present invention is directed to the use of low-dose photodynamic therapy to prevent rejection of donor xenogeneic tissue by modifying the tissue so as to reduce immunogenicity. In this approach, donor tissue is incubated with a suitable photosensitizer and then exposed to light of low intensity. After this treatment, the donor tissue is implanted into a xenogeneic recipient.